Efficient coupling of catalysis and dynamics in the E1 component of Escherichia coli pyruvate dehydrogenase multienzyme complex

Sachin Kale, Gözde Ulas, Jaeyoung Song, Gary W Brudvig, William Furey, Frank Jordan

Research output: Contribution to journalArticle

35 Citations (Scopus)

Abstract

Protein motions are ubiquitous and are intrinsically coupled to catalysis. Their specific roles, however, remain largely elusive. Dynamic loops at the active center of the E1 component of Escherichia coli pyruvate dehydrogenase multienzyme complex are essential for several catalytic functions starting from a predecarboxylation event and culminating in transfer of the acetyl moiety to the E2 component. Monitoring the kinetics of E1 and its loop variants at various solution viscosities, we show that the rate of a chemical step is modulated by loop dynamics. A cysteine-free E1 construct was site-specifically labeled on the inner loop (residues 401-413), and the EPR nitroxide label revealed ligand-induced conformational dynamics of the loop and a slow "open ↔ close" conformational equilibrium in the unliganded state. An 19F NMR label placed at the same residue revealed motion on the millisecond-second time scale and suggested a quantitative correlation of E1 catalysis and loop dynamics for the 200,000-Da protein. Thermodynamic studies revealed that these motions may promote covalent addition of substrate to the enzyme-bound thiamin diphosphate by reducing the free energy of activation. Furthermore, the global dynamics of E1 presumably regulate and streamline the catalytic steps of the overall complex by inducing an entirely entropic (nonmechanical) negative cooperativity with respect to substrate binding at higher temperatures. Our results are consistent with, and reinforce the hypothesis of, coupling of catalysis and regulation with enzyme dynamics and suggest the mechanism by which it is achieved in a key branchpoint enzyme in sugar metabolism.

Original languageEnglish
Pages (from-to)1158-1163
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume105
Issue number4
DOIs
Publication statusPublished - Jan 29 2008

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Multienzyme Complexes
Pyruvate Dehydrogenase Complex
Catalysis
Escherichia coli
Enzymes
Thiamine Pyrophosphate
Thermodynamics
Viscosity
Cysteine
Proteins
Ligands
Temperature

Keywords

  • Coupling of dynamics to catalysis
  • EPR
  • Mobile loop dynamics
  • NMR
  • Pyruvate dehydrogenase

ASJC Scopus subject areas

  • Genetics
  • General

Cite this

Efficient coupling of catalysis and dynamics in the E1 component of Escherichia coli pyruvate dehydrogenase multienzyme complex. / Kale, Sachin; Ulas, Gözde; Song, Jaeyoung; Brudvig, Gary W; Furey, William; Jordan, Frank.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 105, No. 4, 29.01.2008, p. 1158-1163.

Research output: Contribution to journalArticle

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